This application addresses the broad challenge area "(15) Translational Science" and specific challenge topic, "15-HL-102: Develop new therapeutic strategies for heart, lung, and blood diseases based on microRNA technology". The project title is "microRNA Regulation of Human Airway Epithelial Cell Phenotype". microRNAs (miRNAs) play critical roles regulating genomes and genes and are known to be involved in multiple biological processes relevant to lung diseases, such as lung development, lung inflammation, the response to tobacco smoking and lung cancer. However, the expression and function of the majority of miRNAs in critical lung cell types, such as human bronchial epithelial (hBE) cells, remains poorly understood. When hBE cells are grown in vitro at an air-liquid interface (ALI), they recapitulate key aspects of the structure and function of the native epithelium in vivo. The goals of this project are to gain a comprehensive understanding of the miRNA repertoire in ALI hBE cells and its expression under relevant injury/repair conditions. We will then test the function of select, highly regulated miRNAs in this important cell type. The central hypothesis of this proposal is that miRNAs are critical regulators of hBE cell differentiation and the response to injury, that miRNAs become altered in pathologic states, and that modulating miRNAs can change hBE cell phenotype for therapeutic benefit. To test this hypothesis we pose the following specific aims: Aim 1) Develop a comprehensive portrait of miRNA expression in hBE cells during normal differentiation and in a spectrum of relevant injury/repair conditions;and Aim 2) Determine functional consequences of manipulating expression of candidate miRNAs in hBE cells. We will conduct state-of-the art miRNA discovery studies and correlative miRNA and mRNA expression array studies in ALI hBE cells during normal growth and differentiation and under a series of highly relevant injury/repair conditions. We will manipulate miRNA expression as proof-of-concept that modulating miRNAs can alter hBE cell phenotype for therapeutic benefit. These experiments are enabled by a unique combination of resources, expertise and economies of scale and are ambitious, but achievable within the 2-year scope of RFA-OD-09-003. They extend and complement studies in human patient samples by mitigating cell type heterogeneity and sampling concerns. The model system will create a valuable database and will enable evaluation of miRNA function in hBE cells in health and disease, serving as a platform for critical milestone testing of improved treatments for lung diseases based on miRNA. PUBLIC HEALTH RELEVANCE: In this study we will comprehensively determine the miRNA repertoire of a key lung cell type, human bronchial epithelial cells, and will test the function of novel and/or highly regulated miRNAs to change their structure and function. We will create a valuable database and a platform for critical milestone testing of improved treatments for lung diseases that afflict greater than 30 million Americans.